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Abstract

Neuronal mitochondrial dysfunction is a key pathophysiologic mechanism of diabetic peripheral neuropathy (DPN). KU-596 is a small molecule modulator of heat shock protein 90 (Hsp90) that can reverse clinically relevant measures of DPN in diabetic animal models. Mechanistically, drug efficacy requires Hsp70 and correlates with improving mitochondrial bioenergetics (mtBE) in diabetic sensory neurons. The goal of this study was to determine if KU-596 improves mtBE by decreasing glucose-induced oxidative stress in an Hsp70-dependent manner. Sensory neurons were isolated from non-diabetic or diabetic mice wild type (WT) or Hsp70 knockout (Hsp70 KO) mice and treated ex vivo with KU-596 in the presence of low or high glucose concentrations. In diabetic WT and Hsp70 KO neurons, hyperglycemia significantly increased superoxide levels, but KU-596 only decreased superoxide in WT neurons. Similarly, KU-596 significantly improved mtBE in hyperglycemically stressed diabetic WT neurons but did not improve mtBE in diabetic Hsp70 KO under the same conditions. Since manganese superoxide dismutase (MnSOD) is the main mechanism to detoxify mitochondrial superoxide radicals, we determined the cause and effect relationship between improved respiration and decreased oxidative stress by knocking down MnSOD. Downregulating MnSOD in diabetic WT neurons increased hyperglycemia-induced superoxide levels as measured by electron paramagnetic resonance spectroscopy and blocked the ability of KU-596 to enhance mtBE. In diabetic neurons, knockdown of MnSOD increased maximal respiratory capacity (MRC) and this was significantly decreased by mito-TEMPO-H after scavenging overproduced superoxide radicals. Following MnSOD knockdown, KU-596 decreased mitochondrial superoxide but was unable to improve MRC. Overall, this study shows that the Hsp90 inhibitor KU-596 improves mtBE and decreases mitochondrial oxidative stress of diabetic sensory neurons in an Hsp70-dependent manner. However, the drug-induced improvement of mtBE is not necessarily dependent on decreasing mitochondrial oxidative stress and other mechanisms can be modulated by novologue therapy to improve mtBE. This work furthers our mechanistic understanding by which novologue therapy can improve metabolic aspects of neuronal function that contribute to the efficacy of KU-596 for treating diabetic peripheral neuropathy.